This work was supported by the National Natural Science Foundation of China (81870304) to Jun Li and by the Else Kr&#246;ner-Fresenius-Stiftung (Nr. 2017_A28) to Marcus Groettrup.

Inbred 8-10 weeks old male F344 and Lewis rats (200 g to 250 g) were commercially obtained. Allogeneic left kidney transplantation was performed between male F344 and Lewis rats. F344 rats were used as donors and syngeneic recipients, and Lewis rats served as allogeneic recipients. All animal handling procedures were conducted in compliance with guidelines for the Care and Use of Laboratory Animals published by NIH, and all animal experimental protocols were approved by the Animal Care and Use Committee of Chongqing University Cancer Hospital. All&#160;supplies used during surgery, including surgical instruments and solutions, are sterile. A schematic of the protocol is shown in Figure 1.
1. Donor procedure
<ol>
	<li>Induce general anesthesia in the rat by 5% isoflurane inhalation using an induction chamber. Then, subcutaneously inject buprenorphine at 0.1 mg/kg to perform concurrent preemptive analgesia.</li>
	<li>Place the rat on its back and maintain anesthesia with 2% isoflurane inhalation using a facemask over its nose and mouth. Apply eye lubricant onto the eyes to avoid drying of the cornea. Slow respiratory rate and rhythm, the disappearance of corneal reflex, and lack of response to the toe pinch indicate the effectiveness of anesthesia.</li>
	<li>Shave the abdominal hair with an electric razor and sterilize the skin using 0.5% iodine and 70% alcohol.</li>
	<li>Subcutaneously inject 0.5% lidocaine along the midline from the symphysis pubis to subxiphoid for local analgesics, and then incise the abdomen and pull open the incision using a retractor.</li>
	<li>Take out the intestines from the right side of the incision and wrap them up with moistened gauze to prevent them from drying out. Then, expose the left kidney.</li>
	<li>Segregate fat tissues from the left kidney and ureter using cotton swabs, and then dissociate the left renal artery and vein using the micro-forceps under an operating microscope with 20x magnification. Coagulate any bleeding using electrocoagulation, if necessary.</li>
	<li>Perform ligation of the aorta approximately 5 mm above the left renal artery with a 4-0 polyamide monofilament suture. Then, transect the left renal vein distal to the conjunction of the left genital vein and adrenal vein.</li>
	<li>Flush the kidney with ice-cold UW solution supplemented with heparin (100 U/mL) using a 24 G scalp needle from the aorta below the left renal artery until blood fades in color. Warm ischemia time is 5 min on average.</li>
	<li>After transecting the left renal artery approximately 2 mm next to the aorta, dissociate the kidney and ureter with the help of micro-forceps (preserve the peripheral connective tissues to ensure blood supply to the ureter). Then, transect the ureter next to the bladder and preserve the donor left kidney in ice-cold UW solution.</li>
	<li>Sacrifice the donor rat with loss of blood by transecting the aorta and subsequently placing it into a CO2 box to ensure death.</li>
</ol>
2. Recipient procedure
<ol>
	<li>Repeat the procedure described in steps 1.1-1.5 for the recipient rat.</li>
	<li>Dissociate the left kidney and ureter as well as the left renal artery and renal vein of the recipient rat using cotton swabs and micro-forceps under an operating microscope with 20x magnification.</li>
	<li>Clip the left renal artery and renal vein at the root by noninvasive micro-vascular clamps. Ligate the left ureter approximately 2-3 cm below the kidney with an 8-0 polyamide monofilament suture and transect it at ligation.</li>
	<li>Resect the recipient&#39;s native left kidney by transecting the left renal artery 2 mm away from the micro-vascular clamp, and transecting the renal vein proximal to the conjunction of the left genital vein and adrenal vein. Coagulate the adrenal vein using electrocoagulation, if necessary.</li>
	<li>Implant the donor kidney into the left renal fossa of the recipient rat and put ice around the implanted donor kidney. Anastomose the donor&#39;s renal artery and renal vein to the recipient&#39;s renal artery and renal vein in an end-to-end pattern using 10-0 polyamide monofilament sutures under an operating microscope with 45x magnification, as follows.</li>
	<li>Anastomose the renal artery with interrupted sutures.
	<ol>
		<li>Place the stay sutures at 12 and 6 o&#39;clock positions of anastomosis, respectively. Equidistantly suture one side of the anastomosis between the two stay sutures with 2-3 stitches using a 10-0 Polyamide Monofilament suture.</li>
		<li>Turn over the stay sutures and similarly suture the other side of the anastomosis between the two stay sutures with 2-3 stitches.</li>
	</ol>
	</li>
	<li>Anastomose the renal vein with continuous sutures.
	<ol>
		<li>Place the stay sutures at 6 and 12 o&#39;clock positions of anastomosis, respectively. Suture one side of the anastomosis from the 12 o&#39;clock position with 4-5 stitches using the running sutures, and then tie the running suture to the stay suture at the 6 o&#39;clock position with a 10-0 polyamide monofilament suture.</li>
		<li>Turn over the stay sutures and similarly suture the other side of the anastomosis from the 6 o&#39;clock position, and finally tie the running suture to the stay suture at the 12 o&#39;clock position.</li>
	</ol>
	</li>
	<li>Re-perfuse the donor kidney by releasing the noninvasive micro-vascular clamp of the renal vein first; then, identify the bleeding sites and make additional stitches.</li>
	<li>Release the noninvasive micro-vascular clamp of the renal artery, identify the bleeding sites and make additional stitches. Cold ischemia time is 45 min on average.</li>
	<li>Stitch the end of the donor ureter with a 4-0 polyamide monofilament suture as a tow to drag the end through the &#34;tunnel&#34; of the recipient&#39;s bladder under an operating microscope with 20x magnification. Afterward, transect the end of the donor ureter with the suture outside the recipient&#39;s bladder. Let the donor ureter shrink back into the recipient&#39;s bladder.</li>
	<li>Fix the donor ureter with the recipient&#39;s bladder by stitching the adventitia of the donor ureter with the muscle layer of the recipient&#39;s bladder outside at four equidistant positions using an 8-0 polyamide monofilament suture under an operating microscope with 45x magnification.</li>
	<li>Place the intestines back into the abdominal cavity and close the abdominal incision with continuous sutures in the muscle layer first and then the skin layer with a 4-0 polyamide monofilament suture.</li>
	<li>Place the recipient rat on a 37&#176;C heating pad in a dry and clean cage. Wait until the rat recovers from anesthesia.</li>
	<li>Subcutaneously inject buprenorphine (0.05 mg/kg) into the recipient rat every 6 h for 48 h for post-operative analgesia. Intramuscularly inject penicillin (50,000 U/kg) into the recipient rat once a day for 3 days for prevention of infection. The recipient rat is sacrificed by placing it into a CO2 box to observe the chronic rejection of renal allograft after 10 weeks of transplantation.</li>
</ol>

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A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Renal+transplantation+in+rats+with+the+use+of+microsurgical+techniques:+a+new+method.">Renal transplantation in rats with the use of microsurgical techniques: a new method.</a> Surgery. 63 (6), 956-961 (1968).</li><li>Ahmadi, A. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Orthotopic+rat+kidney+transplantation:+A+novel+and+simplified+surgical+approach.">Orthotopic rat kidney transplantation: A novel and simplified surgical approach.</a> Journal of Visualized Experiments: JoVE. (147), e59403 (2019).</li><li>G&#252;nther, E., Walter, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+major+histocompatibility+complex+of+the+rat+(Rattus+norvegicus).">The major histocompatibility complex of the rat (Rattus norvegicus).</a> Immunogenetics. 53 (7), 520-542 (2001).</li><li>Loupy, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Banff+2015+Kidney+Meeting+Report:+Current+challenges+in+rejection+classification+and+prospects+for+adopting+molecular+pathology.">The Banff 2015 Kidney Meeting Report: Current challenges in rejection classification and prospects for adopting molecular pathology.</a> American Journal of Transplantation. 17 (1), 28-41 (2017).</li><li>Fisher, B., Sun, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microvascular+surgical+techniques+in+research,+with+special+reference+to+renal+transplantation+in+the+rat.">Microvascular surgical techniques in research, with special reference to renal transplantation in the rat.</a> Surgery. 58 (5), 904-914 (1965).</li><li>Kamada, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+description+of+cuff+techniques+for+renal+transplantation+in+the+rat.+Use+in+studies+of+tolerance+induction+during+combined+liver+grafting.">A description of cuff techniques for renal transplantation in the rat. Use in studies of tolerance induction during combined liver grafting.</a> Transplantation. 39 (1), 93-95 (1985).</li><li>Grabner, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Non-invasive+imaging+of+acute+allograft+rejection+after+rat+renal+transplantation+using+18F-FDG+PET.">Non-invasive imaging of acute allograft rejection after rat renal transplantation using 18F-FDG PET.</a> Journal of Visualized Experiments: JoVE. (74), e4240 (2013).</li></ol>

The authors have no conflicts of interest to disclose.

Renal transplantation in the rat is a challenging work requiring a high level of micro-surgery techniques and the operation techniques have been optimized several times. From the beginning, Gonzalez et al. implanted the donor kidney into the neck of the recipient and anastomosed the donor ureter to the skin9. However, because of the high incidence of urinary infection and stenosis of the donor ureter, the operation was abandoned in a short time. Subsequently, the implant operation of the donor&#39...

Renal transplantation has become the most effective therapeutic approach for patients with end-stage renal function failure. However, T cell-mediated acute rejection and alloantibody-mediated humoral immune rejection result in the pathologic injury of renal allografts and limit the short-term and long-term survival of patients after kidney transplantation1,2,3. Unfortunately, the effective pharmaceuticals that prevent the rejection of renal allografts are still lacking, because the exact mechanisms of immune and pathologic rejection of renal allografts are not clear. Consequently, the preclinical studies that elucidate the mechanisms of immune and pathologic rejection of renal allografts contribute to finding novel targets and developing relevant effective pharmaceuticals to prevent the rejection of renal allografts and eventually prolong the survival of patients.
Many potential immunological and pathophysiological mechanisms of renal allograft rejection have been proposed recently in rat model studies of orthotopic renal transplantation4,5,6,7,8. These findings propose several novel targets and relevant interfering approaches as promising therapeutics to suppress renal allograft rejection, such as complement regulatory factors and anti-CD59 antibodies6, immunoproteasome, and epoxyketone inhibitors7,8. Thus, rat orthotopic renal transplantation is an ideal preclinical model to investigate the mechanisms of immune rejection and pathologic injury of renal allografts after kidney transplantation.
Rat kidney transplantation has gradually shifted from heterotopic implantation of donor&#39;s kidneys9 to orthotopic renal implantation using end-to-side anastomosis of vessels or using end-to-end anastomosis of the ureter using a cuff method10,11,12. The present study describes detailed protocols of the orthotopic renal transplantation between rats using end-to-end anastomosis to recipients&#39; renal artery and renal vein, and an end-to-side &#34;tunnel&#34; method of ureter-bladder anastomosis, which avoids the ischemia of the lower body and the thrombosis of inferior vena cava and reduces postoperative urine leakage and the twist of the ureter.

<table><tbody><tr><td>&amp;nbsp;10-0 Polyamide Monofilament suture</td><td>B.Braun Medical Inc.</td><td>G0090781</td><td></td></tr><tr><td>&amp;nbsp;4-0 Polyamide Monofilament suture</td><td>B.Braun Medical Inc.</td><td>C1048451</td><td></td></tr><tr><td>&amp;nbsp;8-0 Polyamide Monofilament suture</td><td>B.Braun Medical Inc.</td><td>C2090880</td><td></td></tr><tr><td>Buprenorphine</td><td>US Biological life Sciences</td><td>352004</td><td></td></tr><tr><td>Electrocoagulator</td><td>Electrocoagulator</td><td>ZJ1099</td><td></td></tr><tr><td>F344 and Lewis rats</td><td>Center of Experimental Animals (Tongji Medical College, Huazhong University of Science and Technology, China)</td><td>NA</td><td></td></tr><tr><td>Gauze</td><td>Henan piaoan group Co., LTD</td><td>10210402</td><td></td></tr><tr><td>Heating pad</td><td>Guangzhou Dewei Biological Technology Co., LTD</td><td>DK0032</td><td></td></tr><tr><td>Heparin</td><td>North China Pharmaceutical Co., LTD</td><td>2101131-2</td><td></td></tr><tr><td>Injection syringe (1 ml and 10 ml)</td><td>Shandong weigao group medical polymer Co., LTD</td><td>20211001</td><td></td></tr><tr><td>Isoflurane</td><td>RWD Life Science Co., LTD</td><td>21070201</td><td></td></tr><tr><td>Penicillin G Sodium</td><td>Wuhan HongDe Yuexin pharmatech co.,Ltd</td><td>69-57-8</td><td></td></tr><tr><td>Scalp needle (24 G)</td><td>Hongyu Medical Group</td><td>20183150210</td><td></td></tr><tr><td>Shaver</td><td>Beyotime</td><td>FS600</td><td></td></tr><tr><td>Small animal anesthesia machine</td><td>RWD Life Science</td><td>R500</td><td></td></tr><tr><td>Small Animal Surgery Kit</td><td>Beyotime</td><td>FS500</td><td></td></tr><tr><td>Sodium chloride injection</td><td>Southwest pharmaceutical Co., LTD</td><td>H50021610</td><td></td></tr><tr><td>Surgical operation microscope</td><td>Tiannuoxiang Scientific Instrument Co. , Ltd, Beijing, China</td><td>SZX-6745</td><td></td></tr><tr><td>Swab</td><td>Yubei Medical Materials Co., LTD</td><td>21080274</td><td></td></tr><tr><td>Tape</td><td>Minnesota Mining Manufacturing Medical Equipment (Shanghai) Co., LTD</td><td>1911N68</td><td></td></tr><tr><td>UW solution</td><td>Bristol-Myers Squibb Company</td><td>17HB0002</td><td></td></tr></tbody></table>

a rat orthotopic renal transplantation model for renal allograft rejection

Renal allograft rejection limits the long-term survival of patients after renal transplantation. Rat orthotopic renal transplantation is an essential model to investigate the mechanism of renal allograft rejection in pre-clinical studies and could aid in the development of novel approaches to improve the long-term survival of renal allografts. Donor kidney implantation in rat orthotopic renal transplantation is commonly performed by end-to-side anastomosis to recipients' aorta and inferior vena cava. In this model, the donor's kidney was implanted using end-to-end anastomosis to the recipients' renal artery and renal vein. The donor's ureter was anastomosed to the recipient's bladder in an end-to-side 'tunnel' method. This model contributes to better healing of ureter-bladder anastomosis and increases the recipients' survival by avoiding interference with blood supply and venous reflux of the lower body. This model can be used to investigate the mechanisms of acute and chronic immune and pathologic rejection of renal allografts. Here, the study describes the detailed protocols of this orthotopic renal transplantation between rats.

In this rat orthotopic renal transplantation model, the recipient rats move normally after operation. To observe the chronic rejection of renal allograft, recipient rats are raised for 10 weeks after transplantation, and the total survival rate of recipient rats at this time point is approximately 90%. The major causes of death are bleeding and leakage of urine post-operation. The other major complications include bleeding during the operation, thrombosis in renal vessels, and hydronephrosis, whose respective incidence i...

Watch this Scientific Journal Video about A Rat Orthotopic Renal Transplantation Model for Renal Allograft Rejection at JoVE.com

A Rat Orthotopic Renal Transplantation Model for Renal Allograft Rejection

The rat orthotopic renal transplantation model contributes to investigating the mechanism of renal allograft rejection. The current model increases the recipients' survival without interference with blood supply and venous reflux of the lower body using an end-to-end anastomosis of kidney implantation and an end-to-side "tunnel" method of ureter-bladder anastomosis.

Renal allograft rejection limits the long-term survival of patients after renal transplantation. Rat orthotopic renal transplantation is an essential ...

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Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

3.6K Views.  Chongqing University. The rat orthotopic renal transplantation model contributes to investigating the mechanism of renal allograft rejection. The current model increases the recipients' survival without interference with blood supply and venous reflux of the lower body using an end-to-end anastomosis of kidney implantation and an end-to-side "tunnel" method of ureter-bladder anastomosis.

Video: A Rat Orthotopic Renal Transplantation Model for Renal Allograft Rejection

Orthotopic Aortic Transplantation: A Rat Model to Study the Development of Chronic Vasculopathy

Research models of chronic rejection are essential to investigate pathobiological and pathophysiological processes during the development of transplant vasculopathy (TVP).
The commonly used animal model for cardiovascular chronic rejection studies is the heterotopic heart transplant model performed in laboratory rodents. This model is used widely in experiments since Ono and Lindsey (3) published their technique. To analyze the findings in the blood vessels, the heart has to be sectioned and all vessels have to be measured.
Another method to investigate chronic rejection in cardiovascular questionings is the aortic transplant model (1, 2). In the orthotopic aortic transplant model, the aorta can easily be histologically evaluated (2). The PVG-to-ACI model is especially useful for CAV studies, since acute vascular rejection is not a major confounding factor and Cyclosporin A (CsA) treatment does not prevent the development of CAV, similar to what we find in the clinical setting (4). A7-day period of CsA is required in this model to prevent acute rejection and to achieve long-term survival with the development of TVP.
This model can also be used to investigate acute cellular rejection and media necrosis in xenogeneic models (5).

This video demonstrates the orthotopic aortic transplant model as a simple model to study the development of transplant vasculopathy (TVP) in rats.

Research models of chronic rejection are essential to investigate pathobiological and pathophysiological processes during the development of transplant ...

Non-invasive Imaging of Acute Allograft Rejection after Rat Renal Transplantation Using 18F-FDG PET

The number of patients with end-stage renal disease, and the number of kidney allograft recipients continuously increases. Episodes of acute cellular allograft rejection (AR) are a negative prognostic factor for long-term allograft survival, and its timely diagnosis is crucial for allograft function 1. At present, AR can only be definitely diagnosed by core-needle biopsy, which, as an invasive method, bares significant risk of graft injury or even loss. Moreover, biopsies are not feasible in patients taking anticoagulant drugs and the limited sampling site of this technique may result in false negative results if the AR is focal or patchy. As a consequence, this gave rise to an ongoing search for new AR detection methods, which often has to be done in animals including the use of various transplantation models.
Since the early 60s rat renal transplantation is a well-established experimental method for the examination and analysis of AR 2. We herein present in addition small animal positron emission tomography (PET) using 18F-fluorodeoxyglucose (FDG) to assess AR in an allogeneic uninephrectomized rat renal transplantation model and propose graft FDG-PET imaging as a new option for a non-invasive, specific and early diagnosis of AR also for the human situation 3. Further, this method can be applied for follow-up to improve monitoring of transplant rejection 4.

Non-invasive Imaging of Acute Allograft Rejection after Rat Renal Transplantation Using 18F-FDG PET

We herein present a rat renal transplantation model to non-invasively assess acute allograft rejection using positron emission tomography with 18F-fluorodeoxyglucose.

The number of patients with end-stage renal  disease, and the number of kidney allograft recipients continuously increases.  Episodes of acute cellular ...

Murine Renal Transplantation Procedure

Renal orthotopic transplantation in mice is a technically challenging procedure. Although the first kidney transplants in mice were performed by Russell et al over 30 years ago (1) and refined by Zhang et al years later (2), few people in the world have mastered this procedure. In our laboratory we have successfully performed 1200 orthotopic kidney transplantations with &gt; 90% survival rate. The key points for success include stringent control of reperfusion injury, bleeding and thrombosis, both during the procedure and post-transplantation, and use of 10-0 instead of 11-0 suture for anastomoses.
Post-operative care and treatment of the recipient is extremely important to transplant success and evaluation. All renal graft recipients receive antibiotics in the form of an injection of penicillin immediately post-transplant and sulfatrim in the drinking water continually. Overall animal health is evaluated daily and whole blood creatinine analyses are performed routinely with a portable I-STAT machine to assess graft function.

Renal transplantation in mice is a technically challenging procedure that requires careful post-operative care and treatment for success.

Renal orthotopic transplantation in mice is a technically challenging procedure. Although the first kidney transplants in mice were performed by Russell ...

Biology

Mouse Kidney Transplantation: Models of Allograft Rejection

Rejection of the transplanted kidney in humans is still a major cause of morbidity and mortality. The mouse model of renal transplantation closely replicates both the technical and pathological processes that occur in human renal transplantation. Although mouse models of allogeneic rejection in organs other than the kidney exist, and are more technically feasible, there is evidence that different organs elicit disparate rejection modes and dynamics, for instance the time course of rejection in cardiac and renal allograft differs significantly in certain strain combinations. This model is an attractive tool for many reasons despite its technical challenges. As inbred mouse strain haplotypes are well characterized it is possible to choose donor and recipient combinations to model acute allograft rejection by transplanting across MHC class I and II loci. Conversely by transplanting between strains with similar haplotypes a chronic process can be elicited were the allograft kidney develops interstitial fibrosis and tubular atrophy. We have modified the surgical technique to reduce operating time and improve ease of surgery, however a learning curve still needs to be overcome in order to faithfully replicate the model. This study will provide key points in the surgical procedure and aid the process of establishing this technique.

Here, we present a protocol to study the immunology of rejection. The surgical model presented reports a short operating time and a concise technique. Depending on the donor-recipient strain combination, the transplanted kidney may develop acute cellular rejection or chronic allograft damage, defined by interstitial fibrosis and tubular atrophy.

Rejection of the transplanted kidney in humans is still a major cause of morbidity and mortality. The mouse model of renal transplantation closely ...

Mouse Model of Alloimmune-induced Vascular Rejection and Transplant Arteriosclerosis

Vascular rejection that leads to transplant arteriosclerosis (TA) is the leading representation of chronic heart transplant failure. In TA, the immune system of the recipient causes damage of the arterial wall and dysfunction of endothelial cells and smooth muscle cells. This triggers a pathological repair response that is characterized by intimal thickening and luminal occlusion. Understanding the mechanisms by which the immune system causes vasculature rejection and TA may inform the development of novel ways to manage graft failure. Here, we describe a mouse aortic interposition model that can be used to study the pathogenic mechanisms of vascular rejection and TA. The model involves grafting of an aortic segment from a donor animal into an allogeneic recipient. Rejection of the artery segment involves alloimmune reactions and results in arterial changes that resemble vascular rejection. The basic technical approach we describe can be used with different mouse strains and targeted interventions to answer specific questions related to vascular rejection and TA.

We describe a protocol for aortic interposition grafting in mice. The goal of the protocol is to provide a model with which to study pathological processes and therapeutic strategies relevant to alloimmune reactions in arteries and the resultant arterial changes that contribute to organ transplant failure.

Vascular rejection that leads to transplant arteriosclerosis (TA) is the leading representation of chronic heart transplant failure. In TA, the immune ...

Murine Kidney Transplant Technique

The first mouse kidney transplant technique was published in 19731 by the Russell laboratory. Although it took some years for other labs to become proficient in and utilize this technique, it is now widely used by many laboratories around the world. A significant refinement to the original technique using the donor aorta to form the arterial anastomosis instead of the renal artery was developed and reported in 1993 by Kalina and Mottram 2 with a further advancement coming from the same laboratory in 1999 3. While one can become proficient in this model, a search of the literature reveals that many labs still experience a high proportion of graft loss due to arterial thrombosis. We describe here a technique that was devised in our laboratory that vastly reduces the arterial thrombus reported by others 4,5. This is achieved by forming a heel-and-toe cuff of the donor infra-renal aorta that facilitates a larger anastomosis and straighter blood flow into the kidney.

The goal of this manuscript is to describe the steps required to perform a kidney transplant in a mouse, paying particular attention to the details of the arterial anastomosis.

The first mouse kidney transplant technique was published in 19731 by the Russell laboratory. Although it took some years for other labs to become ...

Orthotopic Hind Limb Transplantation in the Mouse

In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and translational research in the field of transplantation medicine. A vast array of reagents is available exclusively in this setting, including mono- and polyclonal antibodies for both diagnostic and interventional applications. In addition, a vast number of genotyped, inbred, transgenic, and knock out strains allow detailed investigation of the individual contributions of humoral and cellular components to the complex interplay of an immune response and make the mouse the gold standard for immunological research. 
Vascularized Composite Allotransplantation (VCA) delineates a novel field of transplantation using allografts to replace "like with like" in patients suffering traumatic or congenital tissue loss. This surgical methodological protocol shows the use of a non-suture cuff technique for super-microvascular anastomosis in an orthotopic mouse hind limb transplantation model. The model specifically allows for comparison between established paradigms in solid organ transplantation with a novel form of transplants consisting of various different tissue components. Uniquely, this model allows for the transplantation of a viable vascularized bone marrow compartment and niche that have the potential to exert a beneficial effect on the balance of immune acceptance and rejection. This technique provides a tool to investigate alloantigen recognition and allograft rejection and acceptance, as well as enables the pursuit of functional nerve regeneration studies to further advance this novel field of transplantation.

This novel model for orthotopic hind limb transplantation in the mouse, applying a non-suture cuff technique for super-microvascular anastomosis, provides a powerful tool for in&#160;vivo mechanistic immunological research related to vascularized composite allotransplantation (VCA).

In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and ...

Heterotopic Renal Autotransplantation in a Porcine Model: A Step-by-Step Protocol

Kidney transplantation is the treatment of choice for patients suffering from end-stage renal disease. It offers better life expectancy and higher quality of life when compared to dialysis. Although the last few decades have seen major improvements in patient outcomes following kidney transplantation, the increasing shortage of available organs represents a severe problem worldwide. To expand the donor pool, marginal kidney grafts recovered from extended criteria donors (ECD) or donated after circulatory death (DCD) are now accepted for transplantation. To further improve the postoperative outcome of these marginal grafts, research must focus on new therapeutic approaches such as alternative preservation techniques, immunomodulation, gene transfer, and stem cell administration.
Experimental studies in animal models are the final step before newly developed techniques can be translated into clinical practice. Porcine kidney transplantation is an excellent model of human transplantation and allows investigation of novel approaches. The major advantage of the porcine model is its anatomical and physiological similarity to the human body, which facilitates the rapid translation of new findings to clinical trials. This article offers a surgical step-by-step protocol for an autotransplantation model and highlights key factors to ensure experimental success. Adequate pre- and postoperative housing, attentive anesthesia, and consistent surgical techniques result in favorable postoperative outcomes. Resection of the contralateral native kidney provides the opportunity to assess post-transplant graft function. The placement of venous and urinary catheters and the use of metabolic cages allow further detailed evaluation. For long-term follow-up studies and investigation of alternative graft preservation techniques, autotransplantation models are superior to allotransplantation models, as they avoid the confounding bias posed by rejection and immunosuppressive medication.

Porcine models of organ transplantation provide an important platform to study mechanisms of organ preservation. This article describes a heterotopic porcine renal autotransplantation model, which allows investigating new approaches to improve the outcome of transplantation using marginal kidney grafts.

Kidney transplantation is the treatment of choice for patients suffering from end-stage renal disease. It offers better life expectancy and higher quality ...

Orthotopic Rat Kidney Transplantation: A Novel and Simplified Surgical Approach

Kidney transplantation offers increased survival rates and improved quality of life for patients with end-stage renal disease, as compared to any type of renal replacement therapy. Over the past few decades, the rat kidney transplantation model has been used to study the immunological phenomena of rejection and tolerance. This model has become an indispensable tool to test new immunomodulatory pharmaceuticals and regimens prior to proceeding with expensive preclinical large animal studies.
This protocol provides a detailed overview of how to reliably perform orthotopic kidney transplantation in rats. This protocol includes three distinctive steps that increase the probability of success: perfusion of the donor kidney by flushing through the portal vein and the use of a cuff system to anastomose the renal veins and ureters, thereby decreasing cold and warm ischemia times. Using this technique, we have achieved survival rates beyond 6 months with normal serum creatinine in animals with syngeneic or tolerant kidney transplants. Depending on the aim of the study, this model can be modified by pre- or posttransplant treatments to study the acute, chronic, cellular, or antibody-mediated rejection. It is a reproducible, reliable, and cost-effective animal model to study different aspects of kidney transplantation.

The purpose of this manuscript and protocol is to explain and demonstrate in detail the surgical procedure of orthotopic kidney transplantation in rats. This method is simplified to achieve the correct perfusion of the donor kidney and shorten the reperfusion time by using the venous and ureteral cuff anastomosis technique.

Kidney transplantation offers increased survival rates and improved quality of life for patients with end-stage renal disease, as compared to any type of ...

A Simplified Model for Heterotopic Heart Valve Transplantation in Rodents

There is an urgent clinical need for heart valve replacements that can grow in children. Heart valve transplantation is proposed as a new type of transplant with the potential to deliver durable heart valves capable of somatic growth with no requirement for anticoagulation. However, the immunobiology of heart valve transplants remains unexplored, highlighting the need for animal models to study this new type of transplant. Previous rat models for heterotopic aortic valve transplantation into the abdominal aorta have been described, though they are technically challenging and costly. For addressing this challenge, a renal subcapsular transplant model was developed in rodents as a practical and more straightforward method for studying heart valve transplant immunobiology. In this model, a single aortic valve leaflet is harvested and inserted into the renal subcapsular space. The kidney is easily accessible, and the transplanted tissue is securely contained in a subcapsular space that is well vascularized and can accommodate a variety of tissue sizes. Furthermore, because a single rat can provide three donor aortic leaflets and a single kidney can provide multiple sites for transplanted tissue, fewer rats are required for a given study. Here, the transplantation technique is described, providing a significant step forward in studying the transplant immunology of heart valve transplantation.

This protocol describes a simple and efficient method for the transplantation of aortic valve leaflets under the renal capsule to allow for the study of alloreactivity of heart valves.